EARLY PREGNANCY DETECTION
IN DAIRY CATTLE BY BIOPRYN™
ELISA TEST
F. Toth, G. Gabor, Fan Huang and R.G. Sasser
INTRODUCTION
• PSPB is a novel protein that was first reported by scientists
at the University of Idaho. PSPB is located in the giant
binucleate cells of the trophoblastic ectoderm of the
placenta and this indicated that it was either synthesized or
sequestered by those cells. (Figure 1)
• Migration of these cells result the appearance of PSPB in
the maternal circulation (Figure 2).
• PSPB is detectable in serum from 24 to 282 (parturition)
days of gestation and can be applied reliably in dairy cow
herds at 28 to 30 days after breading (Figure 3).
• The PSPB was found in sera of some primiparous cows as
early as 15 days following conception and in sera of most
cows at 24 days following conception.
Figure 1:
The Giant Bi-Nucleated Cells
Caruncle
Cotyledon
Figure 2: Migration of
the Giant Bi-Nucleated Cells
Bi-Nucleated
Cells
Placenta
Uterus
PSPB in
Circulation
PSPB ng/mL
Figure 3:
Variation of PSPB concentration in the
serum
Parturition
Days post breeding
• Complete clearence of the protein from blood may
not occur until 80 to 100 days after parturition. For
this reason testing of blood for a new pregnancy
following parturition may give false positive tests.
• An ELISA test (BioPryn™ by BioTracking
LLC)has been developed for the examination of
the PSPB in cattle 30 days after AI (at least 90
days after the previous parturition) (Figure 4 and
5).
• Decreasing the parturition interval is one of the
most important issues in dairies. Pregnancy
specific protein B (PSPB) is measured for
detection of pregnancy in ruminants.
Figure 4: Biopryn ELISA test – color reaction
Figure 5: Biopryn ELISA test – stop color reaction
OBJECTIVES
The objectives were:
• to verify the BioPRYN™ test is adapted method
for pregnancy detection in cattle.
• to determine the sensitivity of the BioPRYN™
test and compare with the sensitivity of the
ultrasonic examination.
• to define the practicability of PSPB test in
Hungarian field.
• to exam the reliability of PSPB test in monitored
circumstances.
METHODS
Experiment 1
No. of cows
69
No. of farms
1
• The cows were examined by rectal ultrasonography
between 29 and 55 days after AI. Blood samples were
taken at the same time and were assayed for PSPB
with BioPRYN™ when all the samples has been
collected.
Experiment 2
No. of cows
1742
No. of farms
23
• AI’s were carried out in 23 dairies and blood was
drawn from 30 to 36 days after AI and assayed for
early pregnancy by BioPRYN™. To confirm the
pregnancy diagnosis, rectal palpation was done on
the farms at 45-90 days after AI.
Experiment 3
No. of cows
336
No. of farms
1
• In Experiment 3, blood samples were assayed with
BioPRYN™ for early pregnancy detection (30-36
days after AI) in 306 Holstein-Friesian and 30
Hungarian Fleckvieh cows. Confirmation was
done by rectal ultrasonography one day after the
blood collection.
RESULTS
Distribution of Optical Density (OD)
0.6
0.5
0.4
OD
0.3
Series1
0.2
cutoff
0.1
71
66
61
56
51
46
41
36
31
26
21
16
11
6
1
0
Experiment 1
OD values
Mean
Minimum
Maximum
Days After AI
open
pregnant open
pregnant open
pregnant
29 - 35
0.22
n= 20
0.91 0.17
n= 19
0.626 0.31
1.41
36 - 55
0.20
n= 17
0.92 0.16
n= 13
0.704 0.26
1.17
Total
0.21
n= 37
0.92 0.16
n= 32
0.63 0.31
1.41
Cutoff:
0.33
Experiment 2
Number of AI
Correct positive diagnosis
Correct negative diagnosis
False positive diagnosis
False negative diagnosis
Sensitivity
Specificity
Positive predicted value
Negative predicted value
1742
772
638
292
40
95.1
68.6
72.6
94.1
Correct diagnosis: result of PSPB test corresponds with the result of ultrasonic examination. False diagnosis: result of PSPB
test differs from the result of ultrasonic examination. Sensitivity: 100*correct positive diagnosis/correct positive
diagnosis+false negative diagnosis. Specificity: 100*correct negative diagnosis/correct negative diagnosis+false positive
diagnosis. Positive predicted value: 100*correct positive diagnosis/correct positive diagnosis+false positive diagnosis.
Negative predicted value: 100*correct negative diagnosis/correct negative diagnosis+false negative diagnosis.
Experiment 3
Number of AI
Correct positive diagnosis
Correct negative diagnosis
Fals positive diagnosis
Fals negative diagnosis
Sensitivity
Specificity
Positive predicted value
Negative predicted value
336
152
164
20
0
100
89.1
88.4
100
DISCUSSION
• In addition, the more accurate ultrasound
(and done more timely) rather than rectal
palpation was used to confirm pregnancy.
The rate of false positive tests was 5.9% and
positive and negative prediction values were
high (94.0 % and 100 %). The two highly
controlled experiments (1 and 3) show that
BioPRYN™ was an excellent test for
pregnancy detection.
CONCLUSION
• In conclusion, the BioPRYN™ test for PSPB
is a practical and safe system for pregnancy
detection in cattle.